Institute for Clinical Neurobiology, Julius-Maximilians-Universität Würzburg, Versbacherstr. 5, D-97078 Würzburg, Germany.
Department of Chemistry and Pharmacy, Food Chemistry, Emil-Fischer-Center, Friedrich-Alexander-Universität Erlangen-Nürnberg, D-90154 Erlangen, Germany.
Biol Chem. 2018 May 24;399(6):549-563. doi: 10.1515/hsz-2017-0262.
GABAA receptors are ligand-gated anion channels that form pentameric arrangements of various subunits. Positive allosteric modulators of GABAA receptors have been reported as being isolated either from plants or synthesized analogs of known GABAA receptor targeting drugs. Recently, we identified monoterpenes, e.g. myrtenol as a positive allosteric modulator at α1β2 GABAA receptors. Here, along with pharmacophore-based virtual screening studies, we demonstrate that scaffold modifications of myrtenol resulted in the loss of modulatory activity. Two independent approaches, fluorescence-based compound analysis and electrophysiological recordings in whole-cell configurations were used for analysis of transfected cells. C-atoms 1 and 2 of the myrtenol backbone were identified as crucial to preserve positive allosteric potential. A modification at C-atom 2 and lack of the hydroxyl group at C-atom 1 exhibited significantly reduced GABAergic currents at α1β2, α1β2γ, α2β3, α2β3γ and α4β3δ receptors. This effect was independent of the γ2 subunit. A sub-screen with side chain length and volume differences at the C-atom 1 identified two compounds that inhibited GABAergic responses but without receptor subtype specificity. Our combined approach of pharmacophore-based virtual screening and functional readouts reveals that side chain modifications of the bridged six-membered ring structure of myrtenol are crucial for its modulatory potential at GABAA receptors.
GABAA 受体是配体门控阴离子通道，可形成各种亚基的五聚体排列。已经报道了 GABAA 受体的正变构调节剂，它们要么从植物中分离出来，要么是已知 GABAA 受体靶向药物的合成类似物。最近，我们确定了单萜，例如，myrtenol 是 α1β2 GABAA 受体的正变构调节剂。在这里，我们结合基于药效团的虚拟筛选研究，证明了 myrtenol 的骨架修饰导致了调节活性的丧失。荧光化合物分析和全细胞记录两种独立的方法用于转染细胞的分析。myrtenol 骨架的 C-原子 1 和 2 被确定为保持正变构潜能的关键。C-原子 2 的修饰和 C-原子 1 上缺少羟基表现出在 α1β2、α1β2γ、α2β3、α2β3γ 和 α4β3δ 受体上 GABA 能电流的显著降低。这种效应与 γ2 亚基无关。在 C-原子 1 处具有侧链长度和体积差异的子筛选确定了两种抑制 GABA 反应但没有受体亚型特异性的化合物。我们基于药效团的虚拟筛选和功能读出的综合方法表明，myrtenol 的桥接六元环结构的侧链修饰对于其在 GABAA 受体上的调节潜力至关重要。
